Type 1 diabetes mellitus (T1D) is a chronic disease resulting from the autoimmune destruction of insulin-producing pancreatic beta cells. In addition to evolving ideas about the role of beta cells in genetic susceptibility to T1D, there is mounting evidence of diverse beta cell responses to T-cell mediated immune attack. In particular, several recent studies in mice, with some corresponding results in humans, suggest that beta cell stress and dysfunction may not only lead to beta cell death, but could possibly lead to increased beta cell plasticity and/or dedifferentiation. Understanding when and how these responses occur will not only identify potential new biomarkers of T1D, but will facilitate the development of novel therapies to prevent or block disease progression. For this reason, there is growing impetus to characterize the altered molecular and metabolic signature of diabetes-susceptible beta cells at early stages in the disease process. To facilitate the characterization of the cellular and molecular events that are responsible for beta cell dysfunction and destruction early in the T1D process there is a need to develop novel omics technologies that can be used to evaluate available human tissues. Furthermore, given the heterogeneity associated with normal and genetically susceptible T1D islets, it will be critical to develop analytical tools that allow detailed characterization of human pancreatic tissues at single cell and near single cell resolution. In accordance with these goals, the purpose of this grant is to use two powerful and complementary technology platforms ? combFISH and nanoDESI - for in situ single cell resolution omics analyses of human pancreatic tissue (that includes gene and protein expression, metabolite and lipid abundances), supplemented and informed with laser capture microdissection-coupled proteomics and transcriptomics to provide critical insights into T1D.